Micelle assemblies

ABSTRACT

The invention encompasses micelle assemblies, compositions having micelle assemblies, and methods for preparing micelle assemblies and compositions thereof. Also, the invention encompasses compounds of the formula: A-X—Y—Z—R 1  wherein A is a carboxy group or is absent; X is a polyol, Y is —C(═O)—, —C(═S)—, or is absent; Z is O, S, or NH; and R 1  is a polyether, wherein one or more hydroxy groups of the polyol are acylated with a fatty acid residue, wherein the compounds form micelle assemblies. The invention encompasses methods of encapsulating molecules using the compounds of the invention.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. patent application Ser. No.11/006,506 filed on Dec. 7, 2004 and also a continuation under 35 U.S.C.111(a) of PCT/US03/17902, filed on Jun. 6, 2003 and published in Englishon Dec. 18, 2003 as WO 03/103594 A2, which claimed priority under 35U.S.C. 119(e) of U.S. Provisional Application Ser. No. 60/386,920, filedJun. 7, 2002, which applications and publication are incorporated hereinby reference.

GOVERNMENT FUNDING

The invention described herein was made with government support underGrant Number (BES-9983272), awarded by the National Science Foundation.The United States Government has certain rights in the invention.

BACKGROUND OF THE INVENTION

Polymeric micelles are self-assembled amphiphilic block copolymers.These micelles have attracted attention as promising colloidal drugdelivery systems (V. P. Torchilin J. Controlled. Release. 2001, 73, 137;C. Allen, D. et al., Colloids and Surfaces B: Biointerfaces 1999, 16, 3;and H. Otsuka, et al., Current Opinion in Colloid & Interface Science2001, 6, 3). In these colloidal systems, the hydrophobic block typicallyforms the core, essentially a “microcontainer” for a lipophilicpharmaceutical (K. Kataoka, et al., Adv. Drug Delivery Rev. 2001, 47,113). The hydrophilic part forms the outer shell, stabilizing theinterface between the core and the external aqueous environment.Compared to traditional micellar systems, these polymericsurfactant-based drug carriers display apparent advantages such as lowercritical micelle concentration (CMC), improved bioavailability,reduction of toxicity, enhanced permeability across the physiologicalbarriers, and substantial changes in drug biodistribution.

Amphiphilic star-like macromolecules (ASMs) have also been studied fordrug delivery applications. (See, e.g., U.S. patent application Ser. No.09/298,729 filed Apr. 23, 1999; U.S. patent application Ser. No.09/422,295, filed Oct. 21, 1999, and International Patent ApplicationUS00/10050 filed Apr. 18, 2000). The core-shell, amphiphilic structureof ASMs is covalently linked, which makes it thermodynamically stablecompared to conventional micellar systems. Thus, ASM's offer numerousadvantages over conventional micellar systems. Despite these advantages,the use of ASM's is somewhat limited due to the difficulty and costassociated with their preparation. Accordingly, there is a need foradditional micellar systems and reverse micellar systems that possesssome of the advantages associated with the thermodynamic stability ofASM's, but which are easier and less expensive to prepare.

SUMMARY OF THE INVENTION

Applicant has discovered that compounds of formula (I):A-X—Y—Z—R₁  (I)wherein A is a carboxy group or is absent; X is a polyol, Y is —C(═O)—,—C(═S)—, or is absent; Z is O, S or NH; and R₁ is a polyether, whereinone or more hydroxy groups of the polyol are acylated with a fatty acidresidue, will aggregate in a solvent to form micellar structures (e.g.,see FIG. 2).

Additionally, compounds of formula (I) having unsaturated bonds (e.g.,in the fatty acid or polyether groups), can be cross-linked afteraggregate formation to form covalently bound structures (i.e.cross-linked micelles). These aggregates, both cross-linked anduncross-linked, are useful in drug delivery applications, as well as inmany other applications where traditional micelles and ASM's can beapplied. The aggregates formed from compounds of formula (I), bothcross-linked and uncross-linked, can be prepared without much of thedifficulty and cost associated with the preparation of ASM's.

Accordingly, the invention provides a compound of formula (I) asdescribed above. Such compounds of formula (I) are useful intermediatesfor preparing aggregates that can be used in drug delivery applicationsand that can be cross-linked to provide cross-linked micelles that arealso useful in drug delivery applications.

The invention also provides a composition comprising a plurality ofcompounds of formula (I) in a solvent. Such a composition is useful forpreparing aggregates and cross-linked micelles comprising compounds offormula (I).

The invention also provides a composition comprising a plurality ofcompounds of formula (I) in a solvent, wherein the compounds of formula(I) are associated into one or more aggregates.

The invention also provides a composition comprising a cross-linkedmicelle that is formed from plurality of compounds of formula (I) in asolvent, wherein the compounds of formula (I) form one or more aggregatestructures and have been cross-linked to provide the cross-linkedmicelle.

The invention also provides an encapsulate comprising a moleculesurrounded or partially surrounded by an aggregate or a cross-linkedmicelle of the invention.

The invention also provides a method for preparing a cross-linkedmicelle of the invention comprising cross-linking aggregates comprisinga plurality of compounds of formula (I) to provide the cross-linkedmicelle. The invention further provides a method where the aggregatesare formed by combining a plurality of compounds of formula (I) in asolvent.

The invention also provides a method for preparing an encapsulate of theinvention comprising combining a plurality of compounds of formula (I)and a molecule (e.g., a therapeutic agent) in a solvent, and allowingthe compounds of formula (I) to aggregate around the molecule, toprovide the encapsulate (i.e. the molecule surrounded or partiallysurrounded by a plurality of compounds of formula (I)).

The invention also provides a method for preparing an encapsulate of theinvention comprising combining a plurality of compounds of formula (I)and a molecule (e.g., a therapeutic agent) in a solvent, allowing thecompounds of formula (I) to aggregate around the molecule, andcross-linking the compounds of formula (I) to provide the encapsulate(i.e. the molecule encapsulated in a cross-linked micelle).

The invention also provides a composition comprising a solvent, and anaggregate of a plurality of compounds of formula (I) surrounding amolecule (e.g., a therapeutic agent).

The invention also provides a pharmaceutical composition comprising anencapsulate of the invention (i.e. a therapeutic agent surrounded orpartially surrounded by a plurality of compounds of formula (I)); and apharmaceutically acceptable carrier.

The invention also provides a pharmaceutical composition comprising anencapsulate of the invention (i.e. a therapeutic agent encapsulated in across-linked micelle); and a pharmaceutically acceptable carrier.

The invention also provides a method for delivering a therapeutic agentto an animal in need of treatment with the agent comprisingadministering an encapsulate of the invention comprising the agent tothe animal.

The invention also provides intermediates and processes useful forpreparing compounds of formula (I) as described herein.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 shows representative compounds of formula (I) (1-3).

FIG. 2 shows an aggregate of a plurality of compounds of formula (I)prior to cross-linking.

FIG. 3 shows an encapsulate of the invention prior to cross-linking,with a molecule (e.g. “Drug”) encapsulated in an aggregate made up of aplurality of compounds of formula (I).

FIG. 4 shows the encapsulate of FIG. 3 after cross-linking to provide amolecule (e.g. “Drug”) encapsulated in a cross-linked micelle.

FIG. 5 Illustrates the syntheses of representative compounds of formula(I) (6).

FIG. 6 Illustrates the syntheses of representative compounds of formula(I) (7).

FIG. 7 Illustrates the syntheses of representative compounds of formula(I) (10).

FIG. 8 shows an encapsulate of the invention, with a hydrophobicmolecule (e.g. “Drug”) encapsulated in an aggregate made up of aplurality of compounds of formula (I).

DETAILED DESCRIPTION

A is a carboxy group or is absent. When present, A may optionally besubstituted with or attached to a bioactive or therapeutically activemolecule. The bioactive or therapeutically active molecule can be anyknown to one of ordinary skill in the art such as those described below.In a most preferred embodiment, the bioactive or therapeutically activemolecule includes, but is not limited to, vitamin E, sulfonic acids,sulfonates, or salicylic acid.

As used herein the term “polyol” includes straight chain and branchedchain aliphatic groups, as well as mono-cyclic and poly-cyclicaliphatics, which are substituted with two or more hydroxy groups. Apolyol typically has from about 2 carbons to about 20 carbons;preferably, from about 3 carbons to about 12 carbons; and morepreferably from about 4 carbons to about 10 carbons. A polyol alsotypically comprises from about 2 to about 20 hydroxy groups; preferablyfrom about 2 to about 12 hydroxy groups; and more preferably from about2 to about 10 hydroxy groups. A polyol can also optionally besubstituted on a carbon atom with one or more (e.g., 1, 2, or 3) carboxygroups (COOH). These carboxy groups can conveniently be used to link thepolyol to the polyether in a compound of formula (I).

One specific polyol is a mono- or di-carboxyilic acid containing from 1to about 10 carbon atoms and substituted with from 1 to about 10hydroxyl groups. The mono- or di-carboxylic acid may be a straightchained or branched chained aliphatic, or a mono-cyclic or poly-cyclicaliphatic compound. Suitable dicarboxylic acids include mucic acid,malic acid, citromalic acid, alkylmalic acid, hydroxy derivatives ofglutaric acid, and alkyl glutaric acids, tartaric acid, citric acid,hydroxy derivatives of rumadic acid, and the like. Suitablemonocarboxylic acids include 2,2-(bis(hydroxymethyl)propionic acid, andN-[tris(hydroxymethyl)methyl]glycine (tricine).

Another specific polyol is a “saccharide,” which includesmonosaccharides, disaccharides, trisaccharides, polysaccharides andsugar alcohols. The term includes glucose, sucrose, fructose and ribose,as well as deoxy sugars such as deoxyribose and the like. Saccharidederivatives can conveniently be prepared by methods known to the art.Examples of suitable mono-saccharides are xylose, arabinose, and ribose.Examples of di-saccharides are maltose, lactose, and sucrose. Examplesof suitable sugar-alcohols are erythritol and sorbitol.

As used herein, the term polyether includes poly(alkylene oxides) havingbetween about 2 and about 150 repeating units. Typically, thepoly(alkylene oxides) have between about 50 and about 110 repeatingunits. The alkylene oxide units contain from 2 to 10 carbon atoms andmay be straight chained or branched. Preferably, the alkylene oxideunits contain from 2 to 10 carbon atoms. Poly(ethylene glycol) (PEG) ispreferred. Alkoxy-, amino-, carboxy-, and sulfo-terminated poly(alkyleneoxides) are preferred, with methoxy-terminated poly(alkylene oxides)being more preferred.

A preferred polyether has the following structure:R₅—(R₆—O—)_(a)—R₆-Q-

wherein R₅ is a 1 to 20 carbon straight-chain or branched alkyl group,—OH, —OR₇, —NH₂, —NHR₇, —NHR₇R₈, —CO₂H, —SO₃H (sulfo), —CH₂—OH,—CH₂—OR₇, —CH₂O—CH₂—R₇, —CH₂—NH₂, —CH₂—NHR₇, —CH₂—NR₇R₈, —CH₂CO₂H,—CH₂SO₃H, or —O—C(═O)—CH₂—CH₂—C(═O)—O—;

R₆ is a 1 to 10 carbon straight-chain or branched divalent alkylenegroup;

each R₇ and R₈ is independently a 1 to 6 carbon straight-chain orbranched alkylene group;

Q is —O—, —S—, or —NR₇; and

a is an integer from 2 to 150, inclusive.

Another preferred polyether is methoxy terminated polyethylene glycol.

In a compound of formula (I), a poly(alkylene oxide) can be linked tothe polyol, for example, through an ether, thioether, amine, ester,thioester, thioamide, or amide linkage. Preferably, a poly(alkyleneoxide) is linked to the polyol by an ester or amide linkage in acompound of formula (I).

As used herein, the term fatty acid includes fatty acids and fatty oilsas conventionally defined, for example, long-chain aliphatic acids thatare found in natural fats and oils. Fatty acids typically comprise fromabout 2 to about 24 carbon atoms. Preferably, fatty acids comprise fromabout 6 to about 18 carbon atoms. The term “fatty acid” encompassescompounds possessing a straight or branched aliphatic chain and an acidgroup, such as a carboxylate, sulfonate, phosphate, phosphonate, and thelike. The “fatty acid” compounds are capable of “esterifying” or forminga similar chemical linkage with hydroxy groups on the polyol. Examplesof suitable fatty acids include caprylic, capric, lauric, myristic,myristoleic, palmitic, palmitoleic, stearic, oleic, linoleic,eleostearic, arachidic, behenic, erucic, and like acids. Fatty acids canbe derived from suitable naturally occurring or synthetic fatty acids oroils, can be saturated or unsaturated, and can optionally includepositional or geometric isomers. Many fatty acids or oils arecommercially available or can be readily prepared or isolated usingprocedures known to those skilled in the art.

As used herein, the term “aggregate” means a plurality of compounds offormula (I) in a solvent that have organized into an ordered structure,for example, a structure having a hydrophobic core and a surroundinghydrophilic layer, or a structure having a hydrophilic core and asurrounding hydrophobic layer.

As used herein, the term “a plurality of compounds of formula (I)” meansmore than one compound of formula (I). In such a plurality, eachcompound of formula (I) can have the same structure, or the pluralitycan include compounds of formula (I) that have differing structures. Ina preferred embodiment, the term “a plurality of compounds of formula(I)” means more than one compound of formula (I), wherein each of thecompounds of formula (I) has the same structure.

As used herein, a “cross-linked micelle” means an aggregate that hasbeen cross-linked to provide a covalently cross-linked structure.

As used herein, the term “encapsulate” means an aggregate, having amolecule (e.g., a therapeutic agent) surrounded or partially surroundedby a plurality of compounds of formula (I). The term encapsulateincludes structures wherein the compounds of formula (I) have beencross-linked, as well as structures wherein the compounds of formula (I)have not been cross-linked.

As used herein, the term “stabilized encapsulate” means an aggregate,having a molecule (e.g., a therapeutic agent) surrounded or partiallysurrounded by a plurality of compounds of formula (I), whereinunsaturated bonds in the compounds of formula (I) have been cross-linkedto provide a covalently stabilized structure.

As illustrated in FIG. 5, a representative compound of formula (I) canbe prepared by acylating a polyol (4) by reaction with a stoichiometricexcess of a fatty acid chloride to provide acylated polyol (5). Suitableconditions for such an acylation reaction are well known. For example,the reaction can be carried out in the presence of a catalyst, such asZnCl₂, with heating. Suitable acylation conditions are illustrated inthe Examples below. Coupling of (5) with a polyether, for examplethrough an ester linkage, using a suitable coupling agent provides thecompound of formula (I) (6).

When a plurality of compounds of formula (I) are placed in a hydrophilicsolvent (e.g., an aqueous solution comprising water), Applicant hasdiscovered that the compounds of formula (I) will aggregate, with thepolyether portion of the compounds extending into the hydrophilicsolvent, and the fatty acid portions of the compounds forming ahydrophobic core. Such aggregates can solubilize a hydrophobic molecule(e.g., a hydrophobic therapeutic agent) in the aqueous solvent, byencapsulating the hydrophobic molecule in the fatty acid core of theaggregates. The hydrophobic molecule can typically be added to thesolution of the compounds of formula (I) subsequent to aggregation, orthe hydrophobic molecule can be added to the solution of the compoundsof formula (I) prior to aggregation, allowing the aggregates to formaround the molecule. Thus, the aggregates formed from the compounds offormula (I) can function similar to traditional micelles.

Aggregates formed from a plurality of compounds of formula (I) can alsounction like reverse micelles. When a plurality of compounds of formula(I) are placed in a hydrophobic solvent (e.g., an organic solvent likehexanes or methylene chloride), the compounds of formula (I) willaggregate, with the fatty acid portion of the compounds extending intothe hydrophobic solvent, and the polyether portion of the compoundsforming a hydrophilic core. Such aggregates can solubilize a hydrophilicmolecule (e.g., a hydrophilic therapeutic agent) in the organic solvent,by encapsulating the hydrophilic molecule in the polyether core of theaggregates. The hydrophilic molecule can typically be added to thesolution of the compounds of formula (I) subsequent to aggregation, orthe hydrophilic molecule can be added to the solution of the compoundsof formula (I) prior to aggregation, allowing the aggregates to formaround the molecule.

Subsequent to aggregate formation, the compounds of formula (I) thatcomprise unsaturated bonds can be cross-linked to form cross-linkedmicelles, which comprise an aggregate of a plurality of compounds offormula (I) that have been covalently linked. These cross-linkedmicelles can also be used as solubilizing agents for a wide range ofapplications. In some cases, a molecule to be solubilized can be addedto a solution comprising such cross-linked micelles, and the moleculecan locate in the core of the cross-linked micelle, and thus, besolubilized.

Covalently stabilized structures can also be formed by cross-linkingunsaturated bonds in an encapsulate that comprises a molecule to besolubilized. For example, a hydrophobic molecule can be combined with aplurality of compounds of formula (I) in an aqueous solvent such thatthe compounds of formula (I) form aggregates around the hydrophobicmolecule (See FIG. 8). After formation of the aggregates, the compoundsof formula (I) can be cross-linked to provide a stabilized encapsulate.

Typically, the aggregates of the invention have a diameter of from about10 nm to about 1000 nm. The diameters can be measured using any suitableanalytical technique, such as, for example, dynamic light scattering.

Typically, cross-linked micelles of the invention have a number averagemolecular weight between about 1,000 and about 100,000 daltons, whichcan be measured using any suitable analytical technique, such as, forexample, by Gel Permeation Chromatography relative to polystyrenestandards.

Compounds of formula (I) can be used to form aggregates and cross-linkedmicelles that function similar to conventional “micelles” or “reversemicelles.” These aggregates and cross-linked micelles can be used foressentially any application in which conventional micelles or reversemicelles are employed. Examples include drug solubilization, fragranceencapsulation, passive targeting for drug delivery, waste watertreatment, enhanced capillary electrophoresis activation, and inductionof protein crystallization.

Accordingly, as used herein, the term “molecule” includes any compoundthat can be incorporated into an aggregate or a cross-linked micelle asdescribed herein. Typically, “molecules” have solubility properties thatare undesirable and that can be modified by incorporation into anaggregate or a cross-linked micelle of the invention. For example, theterm “molecule” includes therapeutic agents, insecticides, pesticides,herbicides, antiseptics, food additives, fragrances, dyes, diagnosticaids, and the like. Other specific examples of molecules include, butare not limited to:

abietic acid, aceglatone, acenaphthene, acenocoumarol, acetohexamide,acetomeroctol, acetoxolone, acetyldigitoxins, acetylene dibromide,acetylene dichloride, acetylsalicylic acid, alantolactone, aldrin,alexitol sodium, allethrin, allylestrenol, allyl sulfide, alprazolam,aluminum bis(acetylsalicylate), ambucetamide, aminochlothenoxazin,aminoglutethimide, amyl chloride, androstenediol, anethole trithone,anilazine, anthralin, Antimycin A, aplasmomycin, arsenoacetic acid,asiaticoside, astemizole, aurodox, aurothioglycanide, 8-azaguanine,azobenzene;

baicalein, Balsam Peru, Balsam Tolu, barban, baxtrobin, bendazac,bendazol, bendroflumethiazide, benomyl, benzathine, benzestrol,benzodepa, benzoxiquinone, benzphetamine, benzthiazide, benzyl benzoate,benzyl cinnamate, bibrocathol, bifenox, binapacryl, bioresmethrin,bisabolol, bisacodyl, bis(chlorophenoxy)methane, bismuth iodosubgallate,bismuth subgallate, bismuth tannate, Bisphenol A, bithionol, bomyl,bromoisovalerate, bomyl chloride, bomyl isovalerate, bornyl salicylate,brodifacoum, bromethalin, broxyquinoline, bufexamac, butamirate,butethal, buthiobate, butlated hydroxyanisole, butylated hydroxytoluene;

calcium iodostearate, calcium saccharate, calcium stearate, capobenicacid, captan, carbamazepine, carbocloral, carbophenothin, carboquone,carotene, carvacrol, cephaeline, cephalin, chaulmoogfic acid, chenodiol,chitin, chlordane, chlorfenac, chlorfenethol, chlorothalonil,chlorotrianisene, chlorprothixene, chlorquinaldol, chromonar,cilostazol, cinchonidine, citral, clinofibrate, clofazimine, clofibrate,cloflucarban, cionitrate, clopidol, clorindione, cloxazolam, coroxon,corticosterone, coumachlor, coumaphos, coumithoate cresyl acetate,crimidine, crifomate, cuprobam, cyamemazine, cyclandelate, cyclarbamatecymarin, cypennethril;

dapsone, defosfamide, deltamethrin, deoxycorticocosterone acetate,desoximetasone, 10 dextromoramide, diacetazoto, dialifor,diathymosulfone, decapthon, dichlofluani, dichlorophen,dichlorphenamide, dicofol, dicryl, dicmarol, dienestrol,diethylstilbestrol, difenamizole, dihydrocodeinone enol acetate,dihydroergotamine, dihydromorphine, dihydrotachysterol, dimestrol,dimethisterone, dioxathion, diphenane,N-(1,2-diphenylethyl)nicofinamide, dipyrocetyl, disulfamide, dithianone,doxenitoin, drazoxolon, durapatite, edifenphos, emodin, enfenamic acid,erbon, ergocorninine, erythrityl tetranitrate, erythromycin stearate,estriol, ethaverine, ethisterone, ethyl biscomacetate,ethylhydrocupreine, ethyl menthane carboxamide, eugenol, euprocin,exalamide;

febarbamate, fenalamide, fenbendazole, fenipentol, fenitrothion,fenofibrate, fenquizone, fenthion, feprazone, flilpin, filixic acid,floctafenine, fluanisone, flumequine, fluocortin butyl, fluoxymesterone,flurothyl, flutazolam, fumagillin, 5-furfuryl-5-isopropylbarbituficacid, fusafungine, glafenine, glucagon, glutethimide, glybuthiazole,griseofulvin, guaiacol carbonate, guaiacol phosphate, halcinonide,hematoprphyrin, hexachlorophene, hexestrol, hexetidine, hexobarbital,hydrochlorothiazide, hydrocodone, ibuproxam, idebenone, indomethacin,inositol niacinate, iobenzamic acid, iocetamic acid, iodipamide,iomeglamic acid, ipodate, isometheptene, isonoxin,2-isovalerylindane-1,3-dione;

josamycin, 11-ketoprogesterone, laurocapram, 3-O-lauroylpyridoxoldiacetate, lidocaine, lindane, linolenic acid, liothyronine,lucensomycin, mancozeb, mandelic acid, isoamyl ester, mazindol,mebendazole, mebhydroline, mebiquine, melarsoprol, melphalan, menadione,menthyl valerate, mephenoxalone, mephentermine, mephenytoin,meprylcaine, mestanolone, mestranol, mesulfen, metergoline, methallatal,methandriol, methaqualone, 3-methylcholanthrene, methylphenidate,17-methyltestosterone, metipranolol, minaprine, myoral, nafialofos,nafiopidil, naphthalene, 2-naphthyl lactate, 2-(2-naphthyloxy)ethan01,naphthyl salicylate, naproxen, nealbarbital, nemadectin, niclosamide,nicoclonate, nicomorphine, nifuroquine, nifuroxazide, nitracrine,nitromersol, nogalamycin, nordazepam, norethandrolone, norgestrienone;

octavefine, oleandrin, oleic acid, oxazepam, oxazolam, oxeladin,oxwthazaine, oxycodone, oxymesterone, oxyphenistan acetate,paraherquamide, parathion, pemoline, pentaerythritol tetranitrate,pentylphenol, perphenazine, phencarbamide, pheniramine,2-phenyl-6-chlorophenol, phentlmethylbarbituric acid, phenytoin,phosalone, phthalylsulfathiazole, phylloquinone, picadex, pifarnine,piketopfen, piprozolin, pirozadil, plafibride, plaunotol, polaprezinc,polythiazide, probenecid, progesterone, promegestone, propanidid,propargite, propham, proquazone, protionamide, pyrimethamine,pyrimithate, pyrvinium pamoate;

quercetin, quinbolone, quizalofo-ethyl, rafoxanide, rescinnamine,rociverine, ronnel salen, scarlet red, siccmn, simazine, simetfide,sobuzoxane, solan, spironolactone, squalene, stanolone, sucralfate,sulfabenz, sulfaguanole, sulfasalazine, sulfoxide, sulpiride,suxibuzone, talbutal, terguide, testosterone, tetrabromocresol,tetrandrine, thiacetazone, thiocolchicine, thiocftc acid, thioquinox,thioridazine, thiram, thymyl N-isoamylcarbamate, tioxidazole, tioxolone,tocopherol, tolciclate, tolnafiate, triclosan, triflusal, triparanol;

ursolic acid, valinomycin, verapamil, vinblastine, vitamin A, vitamin D,vitamin E, xenbucin, xylazine, zaltoprofen, and zearalenone.

The aggregates and cross-linked micelles the invention are particularlyuseful for solubilizing hydrophobic molecules, particularly therapeuticagents that are hydrophobic in nature. Thus, according to one embodimentof the present invention, a therapeutic agent is encapsulated bycombining the agent and a plurality of compounds of formula (I) in asolvent, such as water. After aggregates have formed (which can bedetermined, for example, by using dynamic light scattering, fluorescencespectroscopy, surface tension, or a combination thereof) the compoundsof formula (I) are cross-linked to provide an encapsulate of theinvention wherein the therapeutic agent is encapsulated in across-linked micelle.

The present invention contemplates the use of encapsulated hydrophobicmolecules at concentrations as high as 1 M and greater, up to 10⁶ M. Atthe same time, another advantage of the present invention is thethermodynamic stability of the polymers, which permit the formation oflow concentration stable aqueous solutions of the polymer encapsulates,far below the CMC's of conventional surfactants. Stable aqueoussolutions with concentrations of 10⁻⁸ and greater are expected to havethe greatest commercial utility. According to the invention,encapsulates are believed to form at concentrations below the presentlyavailable limits of detection, i.e., below 10⁻¹⁰ M.

The encapsulates of the invention that comprise a therapeutic agent canbe formulated as pharmaceutical compositions and administered to amammalian host, such as a human patient in a variety of forms adapted tothe chosen route of administration, i.e., orally or parenterally, byintravenous, intramuscular, topical or subcutaneous routes.

Thus, the encapsulates of the invention may be systemicallyadministered, e.g., orally, in combination with a pharmaceuticallyacceptable vehicle such as an inert diluent. They may be incorporateddirectly with the food of the patient's diet. For oral therapeuticadministration, the encapsulates of the invention may be used in theform of elixirs, syrups, and the like.

The compositions may also contain a sweetening agent such as sucrose,fructose, lactose or aspartame or a flavoring agent such as peppermint,oil of wintergreen, or cherry flavoring may be added. A syrup or elixirmay contain the active compound, sucrose or fructose as a sweeteningagent, methyl and propylparabens as preservatives, a dye and flavoringsuch as cherry or orange flavor. Of course, any material used inpreparing any unit dosage form should be pharmaceutically acceptable andsubstantially non-toxic in the amounts employed. In addition, theencapsulates of the invention may be incorporated into sustained-releasepreparations and devices.

The encapsulates of the invention may also be administered intravenouslyor intraperitoneally by infusion or injection. Solutions of theencapsulates can be prepared, for example, in water. Under ordinaryconditions of storage and use, these preparations may contain apreservative to prevent the growth of microorganisms.

The pharmaceutical dosage forms suitable for injection or infusionshould be sterile, fluid and stable under the conditions of manufactureand storage. The prevention of the action of microorganisms can bebrought about by various antibacterial and antifungal agents, forexample, parabens, chlorobutanol, phenol, sorbic acid, thimerosal, andthe like. In many cases, it will be preferable to include isotonicagents, for example, sugars, buffers or sodium chloride.

Sterile injectable solutions are prepared by incorporating theencapsulates of the invention in the required amount in the appropriatesolvent with various of the other ingredients enumerated above, asrequired, followed by sterilization.

Encapsulation of molecules according to the invention modifiestransdermal delivery of the molecule. Absorption through the skin can beincreased or decreased by a factor of up to about 1000. Thus, thepharmaceutical dosage forms of present invention include dosage formssuitable for transdermal delivery, which, in addition to aqueoussolutions, also include aqueous gels. The dosage form may be applieddirectly to the skin as a lotion, cream or salve, or a transdermal drugdelivery device such as a transdermal patch may be employed, in whichthe encapsulated molecule is retained in the active agent reservoir ofthe patch.

The dose and method of administration will vary from animal to animaland be dependent upon such factors as the type of animal being treated,its sex, weight, diet, concurrent medication, overall clinicalcondition, the particular therapeutic agent employed, the specific usefor which the agent is employed, and other factors which those skilledin the relevant field will recognize.

Therapeutically effective dosages may be determined by either in vitroor in vivo methods. For each particular dosage form of the presentinvention, individual determinations may be made to determine theoptimal dosage required. The range of therapeutically effective dosageswill naturally be influenced by the route of administration, thetherapeutic objectives, and the condition of the patient. Thedetermination of effective dosage levels, that is, the dosage levelsnecessary to achieve the desired result, will be within the ambit of oneskilled in the art. Typically, applications of agent are commenced atlower dosage levels, with dosage levels being increased until thedesired effect is achieved.

A typical dosage might range from about 0.001 mg to about 1,000 mg oftherapeutic agent, per kg of animal weight. Preferred dosages range fromabout 0.01 mg/kg to about 100 mg/kg, and more preferably from about 0.10mg/kg to about 20 mg/kg. Advantageously, the dosage forms of thisinvention may administered several times daily, and other dosageregimens may also be useful.

According to the invention, aggregate or cross-linked micelledegradation is not a prerequisite for release of the molecule (e.g. thetherapeutic agent).

The compounds of Formula (I), aggregates, encapsulates and cross-linkedmicelles of the invention may also be used as thickening agents,lubricants, detergents surfactants, plasticizers and anti-foulingagents. The compounds of Formula (I), aggregates, encapsulates andcross-linked micelles of the invention may be used as an emulsifying,dispersing or stabilizing agent for dyes, cosmetics, pigment andpharmaceutical products. The compounds of Formula (I), aggregatesencapsulates and cross-linked micelles of the invention are particularlyuseful as an, emulsifying, dispersing or stabilizing agent in the dyeingof textiles and for encapsulating dyes, fragrances, or both forcosmetics. The compounds of Formula (I), aggregates, encapsulates andcross-linked micelles of the invention are useful as lubricants and as athickening agents for paints. The compounds of Formula (I), aggregates,encapsulates and cross-linked micelles of the invention may also beemployed as an emulsifying, dispersing or stabilizing agent forcomponents of photographic compositions and developers.

For therapeutic applications, the preferred cross-linked micelles andaggregates of the invention hydrolyze into components known to bebiocompatible, i.e., sugars, fatty acids, amino acids and poly(ethyleneglycol). This also results in low cytotoxicity of the polymer and itshydrolysis products. The poly(alkylene oxide) units decrease theimmunogenicity of the encapsulate, enabling the hydrophobic molecules toevade the body's immune system, thereby increasing the circulation timeof the hydrophobic molecule. This allows for effective treatment withreduced quantities of the hydrophobic molecule, which, together with thedecreased immunogenicity, prevents or reduces the severity of incidentsof toxic side effects.

The following non-limiting examples set forth hereinbelow illustratecertain aspects of the invention. All parts and percentages are byweight unless otherwise noted and all temperatures are in degreesCelsius.

All PEG's were obtained from Shearwater Polymers (Birmingham, Ala.) andused without further purification. All other chemicals were obtainedfrom Aldrich (Milwaukee, Wis.), and used without further purification.Analytical grade solvents were used for all the reactions. Methylenechloride, tetrahydrofuran (THF), triethylamine (TEA) anddimethylsulfoxide (DMSO) were distilled. 4-(dimethylamino) pyridiniump-toluenesulfonate (DPTS) was prepared as described by J. S. Moore, S.I. Stupp Macromolecules 1990, 23, 65. ¹H-NMR and spectra were recordedon a Varian 200 MHz or 400 MHz spectrometer. Samples (˜5-10 mg/ml) weredissolved in CDCl₃ or THF-d₄, with the solvent used as an internalreference. IR spectra were recorded on a Mattson Seriesspectrophotometer by solvent casting samples onto a KBr pellet. Thermalanalysis data were determined on a Perkin-Elmer Pyris 1 DSC system,samples (˜10 mg) were heated under dry nitrogen gas. Data were collectedat heating and cooling rates of 5° C./min. Gel permeation chromatography(GPC) was performed on a Perkin-Elmer Series 200 LC system. Dynamiclaser scattering (DSL) measurements were carried on NICOMP particlesizing systems.

EXAMPLES Example 1 Compound (10) (FIG. 7, R=Decyl)

mPEG5k (5.0 g, 1.0 mmol) was dehydrated by azeotropic distillation intoluene (30 ml), and the toluene was removed under vacuum. Compound (9)(1.4 g, 3.0 mmol) and DPTS (0.32 g, 1.0 mmol) in methylene chloride (30ml) were added at room temperature. After 10 minutes flushing withnitrogen, 1.0 M DCC in methylene chloride (3.0 ml) was added dropwise.After 3 days, the DCC side product (dicyclohexylurea) was removed bysuction filtration. The filtrate was washed with 30 ml portions of brine(3), dried over anhydrous sodium sulfate and evaporated to dryness Thecrude product was purified by precipitation into diethyl ether frommethanol. The title compound was obtained as a white waxy solid (5.3 g),yield: 95%.

The intermediate Compound (9) was prepared as follows.

a. Compound (8). bis(Hydroxymethyl)propionic acid (DMPA, 2.0 g, 15 mmol)was dissolved into pyridine (25 ml). After 10 minutes flushing withnitrogen in ice/water bath, 10-undecenoyl chloride (7.4 ml, 34 mmol) wasadded dropwise. After 10 hours, the reaction mixture was poured into 0.1N HCl solutions (300 ml) and stirred for 10 minutes. Ethyl ether (100ml) was used to extract the product. The extraction was washed with 50ml portions of brine (5), dried over anhydrous sodium sulfate, andevaporated to dryness. The crude product was purified by chromatographyusing petroleum ether:ethyl acetate (80:20) as eluent. Compound (9) wasobtained as a colorless oil (5.0 g), yield: 72%.

Examples 2-4 detail the preparation of other acylated polyols that canbe coupled with a polyether to provide compounds of formula (I).

Example 2 FIG. 5, Compound (5), R₂=Ethyl

To a neat mixture of mucic acid (4.2 g, 20 mmol) and propionyl chloride(18 ml, 200 mmol) was added ZnCl₂ (0.28 g, 2.0 mmol). The reactionmixture was heated at reflux temperature for three hours. After cooling,diethyl ether (20 ml) was added to the reaction mixture and the solutionpoured onto ice chips (approximately 100 g) with stirring. Additionaldiethyl ether (80 ml) was added to the mixture and stirring continuedfor 30 minutes more. The ether portion was separated, washed with waterto a neutral pH, dried over anhydrous Na₂SO₄ and evaporated to dryness.The crude product was purified by recrystallization from a cosolventsystem of diethyl ether and methylene chloride, collected by vacuumfiltration, washed by ice cold methylene chloride and dried at 105 EC(12 hours) to 15 constant weight. The title compound was obtained as awhite solid having a T_(m) of 196EC, 56% yield.

Example 3 FIG. 5, Compound (5), R₂=Propyl

Mucic acid hexyl ester was prepared as in Example 2, substitutingcaproyl chloride for propionyl chloride. The title compound was obtainedas a white solid having a T_(m) of 171EC, yield of 68%.

Example 4 FIG. 5, Compound (5), R₂=Undecyl

Mucic acid lauryl ester was prepared as in Example 2, substitutinglauryl chloride for propionyl chloride. The title compound was obtainedas a white solid having a T_(m) of 145EC, yield of 65%.

All publications, patents, and patent documents are incorporated byreference herein, as though individually incorporated by reference. Theinvention has been described with reference to various specific andpreferred embodiments and techniques. However, it should be understoodthat many variations and modifications may be made while remainingwithin the spirit and scope of the invention.

1. A compound of formula (I):A-X—Y—Z—R₁  (I) wherein A is a carboxy group; X is a straight chain orbranched chain aliphatic group containing 2 carbons to about 20 carbonswherein the aliphatic group is substituted with 2 to about 20 hydroxygroups; Y is —C(═O)—, —C(═S)—, or is absent; Z is O, S or NH; and R₁ isa polyether, wherein one or more hydroxy groups of X are acylated with afatty acid residue.
 2. The compound of claim 1 wherein the polyether islinked to X through an ester, thioester, or amide linkage.
 3. Thecompound of claim 1 wherein the fatty acids comprise from about 2 toabout 24 carbon atoms.
 4. The compound of claim 1 wherein the polyetheris a poly(alkylene oxide) having between about 2 and about 150 repeatingunits, the poly(alkylene oxide) is terminated with hydroxy-, alkoxy-,amino-, carboxy-, or sulfo-group and the polyether is linked to Xthrough an ester or amide linkage.
 5. The compound of claim 4 wherein Xhas from about 3 carbons to about 12 carbons wherein the carbons aresubstituted with 2 to about 12 hydroxy groups and the poly(alkyleneoxide) is terminated with a hydroxy-, amino-, carboxy- or sulfo-group.6. The compound of claim 4 wherein the fatty acid is lauric, myristic,myristoleic, palmitic, palmitoleic, stearic, oleic, linoleic, arachidic,behenic, lignoceric, eleostearic or erucic acid, or a mixture thereof.7. The compound of claim 6 wherein the alkylene oxide units contain from2 to 4 carbon atoms which are straight chained or branched, and X hasfrom about 4 carbons to about 10 carbons wherein the carbons aresubstituted with 2 to about 10 hydroxy groups.
 8. The compound of claim5 wherein the alkylene oxide is polyethylene glycol with from 50 to 110repeating units.
 9. The compound of claim 1 wherein the polyether is amethoxy terminated polyethylene glycol.
 10. The compound of claim 1wherein X has from about 4 carbons to about 10 carbons wherein thecarbons are substituted with 2 to about 10 hydroxy groups.
 11. Thecompound of claim 1 wherein X is substituted with one or more carboxygroups.
 12. A composition comprising a plurality of compounds of claim 1in a solvent, wherein the compounds form one or more aggregatestructures.
 13. The composition of claim 12 wherein the solventcomprises an organic solvent.
 14. The composition of claim 12 whereinthe solvent comprises water.
 15. The composition of claim 12 wherein thesolvent is water.
 16. A composition comprising a solvent and across-linked micelle comprising a plurality of compounds of claim 1,wherein the compounds form one or more aggregate structures that arecross-linked to provide the cross-linked micelle.
 17. A method forpreparing an aggregate structure of compounds of claim 1 comprisingcombining a plurality of compounds of formula (I) in a solvent; andallowing them to form the aggregate structure.
 18. A method forpreparing a cross-linked micelle comprising combining a plurality ofcompounds of claim 1 in a solvent; allowing the compounds to formaggregate structures; and cross-linking the compounds to provide thecross-linked micelle.
 19. An encapsulate comprising a moleculesurrounded or partially surrounded by an aggregate of a plurality ofcompounds of claim
 1. 20. The encapsulate of claim 19 wherein themolecule is a therapeutic agent.
 21. An encapsulate comprising amolecule in the cross-linked micelle of claim
 16. 22. A method forpreparing an encapsulate comprising combining a plurality of compoundsof claim 1 and a molecule in a solvent; and allowing the compounds toaggregate around the molecule, to provide the encapsulate.
 23. A methodfor preparing an encapsulate comprising combining a plurality ofcompounds of claim 1 and a molecule, in a solvent; allowing thecompounds to aggregate around the molecule; and cross-linking thecompounds to provide the encapsulate.
 24. A method for delivering atherapeutic agent to an animal in need of treatment with the agentcomprising administering an encapsulate of claim 20 to the animal. 25.The compound of claim 1 wherein the polyether is linked to X through anester or amide linkage.
 26. The compound of claim 1 wherein thepolyether is linked to X through an ester linkage.
 27. The compound ofclaim 1 wherein the fatty acids comprise from about 6 to about 18 carbonatoms.
 28. The compound of claim 1 wherein the fatty acids comprise fromabout 2 to about 24 carbon atoms and include at least one unsaturatedbond.
 29. The compound of claim 1 wherein X has from about 3 carbons toabout 12 carbons wherein the carbons are substituted with 2 to about 12hydroxy groups.
 30. The compound of claim 1 wherein the polyethercomprises alkylene oxide units containing from 2 to 10 carbon atomswhich are straight chained or branched.
 31. The compound of claim 1wherein the polyether comprises alkylene oxide units containing from 2to 4 carbon atoms which are straight chained or branched.